Multiplexed Proteome Dynamics Profiling Reveals Mechanisms Controlling Protein Homeostasis.

Protein degradation plays important roles in biological processes and is tightly regulated. Further, targeted proteolysis is an emerging research tool and therapeutic strategy. However, proteome-wide technologies to investigate the causes and consequences of protein degradation in biological systems are lacking. We developed "multiplexed proteome dynamics profiling" (mPDP), a mass-spectrometry-based approach combining dynamic-SILAC labeling with isobaric mass tagging for multiplexed analysis of protein degradation and synthesis. In three proof-of-concept studies, we uncover different responses induced by the bromodomain inhibitor JQ1 versus a JQ1 proteolysis targeting chimera; we elucidate distinct modes of action of estrogen receptor modulators; and we comprehensively classify HSP90 clients based on their requirement for HSP90 constitutively or during synthesis, demonstrating that constitutive HSP90 clients have lower thermal stability than non-clients, have higher affinity for the chaperone, vary between cell types, and change upon external stimuli. These findings highlight the potential of mPDP to identify dynamically controlled degradation mechanisms in cellular systems.

Structural and functional investigation of ABC transporter STE6-2p from Pichia pastoris reveals unexpected interaction with sterol molecules.

Adenosine triphosphate (ATP)-binding cassette (ABC) transporters are multidomain transmembrane proteins, which facilitate the transport of various substances across cell membranes using energy derived from ATP hydrolysis. They are important drug targets since they mediate decreased drug susceptibility during pharmacological treatments. For the methylotrophic yeast Pichia pastoris, a model organism that is a widely used host for protein expression, the role and function of its ABC transporters is unexplored. In this work, we investigated the Pichia ABC-B transporter STE6-2p. Functional investigations revealed that STE6-2p is capable of transporting rhodamines in vivo and is active in the presence of verapamil and triazoles in vitro. A phylogenetic analysis displays homology among multidrug resistance (MDR) transporters from pathogenic fungi to human ABC-B transporters. Further, we present high-resolution single-particle electron cryomicroscopy structures of an ABC transporter from P. pastoris in the apo conformation (3.1 Å) and in complex with verapamil and adenylyl imidodiphosphate (AMP-PNP) (3.2 Å). An unknown density between transmembrane helices 4, 5, and 6 in both structures suggests the presence of a sterol-binding site of unknown function.

Multiplexed cellular profiling identifies an organoselenium compound as an inhibitor of CRM1-mediated nuclear export.

Chromosomal region maintenance 1 (CRM1 also known as Xpo1 and exportin-1) is the receptor for the nuclear export controlling the intracellular localization and function of many cellular and viral proteins that play a crucial role in viral infections and cancer. The inhibition of CRM1 has emerged as a promising therapeutic approach to interfere with the lifecycle of many viruses, for the treatment of cancer, and to overcome therapy resistance. Recently, selinexor has been approved as the first CRM1 inhibitor for the treatment of multiple myeloma, providing proof of concept for this therapeutic option with a new mode of action. However, selinexor is associated with dose-limiting toxicity and hence, the discovery of alternative small molecule leads that could be developed as less toxic anticancer and antiviral therapeutics will have a significant impact in the clinic. Here, we report a CRM1 inhibitor discovery platform. The development of this platform includes reporter cell lines that monitor CRM1 activity by using red fluorescent protein or green fluorescent protein-labeled HIV-1 Rev protein with a strong heterologous nuclear export signal. Simultaneously, the intracellular localization of other proteins, to be interrogated for their capacity to undergo CRM1-mediated export, can be followed by co-culturing stable cell lines expressing fluorescent fusion proteins. We used this platform to interrogate the mode of nuclear export of several proteins, including PDK1, p110α, STAT5A, FOXO1, 3, 4 and TRIB2, and to screen a compound collection. We show that while p110α partially relies on CRM1-dependent nuclear export, TRIB2 is exported from the nucleus in a CRM1-independent manner. Compound screening revealed the striking activity of an organoselenium compound on the CRM1 nuclear export receptor.

Metabolism of a fungicide propiconazole by Cunninghamella elegans ATCC36112.

The metabolic breakdown of propiconazole by fungi was examined, and it was found that the microbial model (Cunninghamella elegans ATCC36112) efficiently degrades the triazole fungicide propiconazole through the action of cytochrome P450. This enzyme primarily facilitates the oxidation and hydrolysis processes involved in phase I metabolism. We observed major metabolites indicating hydroxylation/oxidation of propyl groups of propiconazole. Around 98% of propiconazole underwent degradation within a span of 3 days post-treatment, leading to the accumulation of five metabolites (M1-M5). The experiments started with a preliminary identification of propiconazole and its metabolites using GC-MS. The identified metabolites were then separated and identified by in-depth analysis using preparative UHPLC and MS/MS. The metabolites of propiconazole are M1 (CGA-118245), M2(CGA-118244), M3(CGA-136735), M4(GB-XLIII-42-1), and M5(SYN-542636). To further investigate the role of key enzymes in potential fungi, we treated the culture medium with piperonyl butoxide (PB) and methimazole (MZ), and then examined the kinetic responses of propiconazole and its metabolites. The results indicated a significant reduction in the metabolism rate of propiconazole in the medium treated with PB, while methimazole showed weaker inhibitory effects on the metabolism of propiconazole in the fungus C. elegans.

Assessing the Toxicity of Benzotriazole Ultraviolet Stabilizers to Fishes: Insights into Aryl Hydrocarbon Receptor-Mediated Effects.

Benzotriazole ultraviolet stabilizers (BUVSs) are chemicals used to mitigate UV-induced damage to manufactured goods. Their presence in aquatic environments and biota raises concerns, as certain BUVSs activate the aryl hydrocarbon receptor (AhR), which is linked to adverse effects in fish. However, potencies of BUVSs as AhR agonists and species sensitivities to AhR activation are poorly understood. This study evaluated the toxicity of three BUVSs using embryotoxicity assays. Zebrafish (Danio rerio) embryos exposed to BUVSs by microinjection suffered dose-dependent increases in mortality, with LD50 values of 4772, 11 608, and 56 292 ng/g-egg for UV-P, UV-9, and UV-090, respectively. The potencies and species sensitivities to AhR2 activation by BUVSs were assessed using a luciferase reporter gene assay with COS-7 cells transfected with the AhR2 of zebrafish and eight other fishes. The rank order of potency for activation of the AhR2 from all nine species was UV-P > UV-9 > UV-090. However, AhR2s among species differed in sensitivities to activation by up to 100-fold. An approximate reversed rank order of species sensitivity was observed compared to the rank order of sensitivity to 2,3,7,8-tetrachlorodibenzo[p]dioxin, the prototypical AhR agonist. Despite this, a pre-existing quantitative adverse outcome pathway linking AhR activation to embryo lethality could predict embryotoxicities of BUVSs in zebrafish.

Discovery of 1,2,3-triazole incorporated indole-piperazines as potent antitubercular agents: Design, synthesis, in vitro biological evaluation, molecular docking and ADME studies.

In this report, a library consisting of three sets of indole-piperazine derivatives was designed through the molecular hybridization approach. In total, fifty new hybrid compounds (T1-T50) were synthesized and screened for antitubercular activity against Mycobacterium tuberculosis H37Rv strain (ATCC-27294). Five (T36, T43, T44, T48 and T49) among fifty compounds exhibited significant inhibitory potency with the MIC of 1.6 µg/mL, which is twofold more potent than the standard first-line TB drug Pyrazinamide and equipotent with Isoniazid. N-1,2,3-triazolyl indole-piperazine derivatives displayed improved inhibition activity as compared to the simple and N-benzyl indole-piperazine derivatives. In addition, the observed activity profile of indole-piperazines was similar to standard anti-TB drugs (isoniazid and pyrazinamide) against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa strains, demonstrating the compounds' selectivity towards the Mycobacterium tuberculosis H37Rv strain. All the active anti-TB compounds are proved to be non-toxic (with IC50 > 300 µg/mL) as verified through the toxicity evaluation against VERO cell lines. Additionally, molecular docking studies against two target enzymes (Inh A and CYP121) were performed to validate the activity profile of indole-piperazine derivatives. Further, in silico-ADME prediction and pharmacokinetic parameters indicated that these compounds have good oral bioavailability.

Computational simulations uncover enantioselective metabolism of chiral triazole fungicides by human CYP450 enzymes: A case study of tebuconazole.

Tebuconazole (TEB), a prominent chiral triazole fungicide, has been extensively utilized for plant pathogen control globally. Despite experimental evidence of TEB metabolism in mammals, the enantioselectivity in the biotransformation of R- and S-TEB enantiomers by specific CYP450s remains elusive. In this work, integrated in silico simulations were employed to unveil the binding interactions and enantioselective metabolic fate of TEB enantiomers within human CYP1A2, 2B6, 2E1, and 3A4. Molecular dynamics (MD) simulations clearly delineated the binding specificity of R- and S-TEB to the four CYP450s, crucially determining their differences in metabolic activity and enantioselectivity. The primary driving force for robust ligand binding was identified as van der Waals interactions with CYP450s, particularly involving the hydrophobic residues. Mechanistic insights derived from quantum mechanics/molecular mechanics (QM/MM) calculations established C2-methyl hydroxylation as the predominant route of R-/S-TEB metabolism, while C6-hydroxylation and triazol epoxidation were deemed kinetically infeasible pathways. Specifically, the resulting hydroxy-R-TEB metabolite primarily originates from R-TEB biotransformation by 1A2, 2E1 and 3A4, whereas hydroxy-S-TEB is preferentially produced by 2B6. These findings significantly contribute to our comprehension of the binding specificity and enantioselective metabolic fate of chiral TEB by CYP450s, potentially informing further research on human health risk assessment associated with TEB exposure.

Mycoremediation of the novel fungicide ametoctradin by different agricultural soils and accelerated degradation utilizing selected fungal strains.

Accelerating safety assessments for novel agrochemicals is imperative, advocating for in vitro setups to present pesticide biodegradation by soil microbiota before field studies. This approach enables metabolic profile generation in a controlled laboratory environment eliminating extrinsic factors. In the current study, ten different soil samples were utilized to check their capability to degrade Ametoctradin by their microbiota. Furthermore, five different fungal strains (Aspergillus niger, Aspergillus flavus, Aspergillus fumigatus, Lasiodiplodia theobromae, and Penicillium chrysogenum) were utilized to degrade Ametoctradin in aqueous media. A degradation pathway was established using the metabolic patterns created during the biodegradation of Ametoctradin. In contrast to 47% degradation (T1/2 of 34 days) when Ametoctradin was left in the soil samples, the fungal strain Aspergillus fumigatus demonstrated 71% degradation of parent Ametoctradin with a half-life (T1/2) of 16 days. In conclusion, soil rich in microorganisms effectively cleans Ametoctradin-contaminated areas while Fungi have also been shown to be an effective, affordable, and promising way to remove Ametoctradin from the environment.

Synthesis and Pharmacological Evaluation of Novel Triazole-Pyrimidine Hybrids as Potential Neuroprotective and Anti-neuroinflammatory Agents.

OBJECTIVE: Neuroprotection is a precise target for the treatment of neurodegenerative diseases, ischemic stroke, and traumatic brain injury. Pyrimidine and its derivatives have been proven to use antiviral, anticancer, antioxidant, and antimicrobial activity prompting us to study the neuroprotection and anti-inflammatory activity of the triazole-pyrimidine hybrid on human microglia and neuronal cell model. METHODS: A series of novel triazole-pyrimidine-based compounds were designed, synthesized and characterized by mass spectra, 1HNMR, 13CNMR, and a single X-Ray diffraction analysis. Further, the neuroprotective, anti-neuroinflammatory activity was evaluated by cell viability assay (MTT), Elisa, qRT-PCR, western blotting, and molecular docking. RESULTS: The molecular results revealed that triazole-pyrimidine hybrid compounds have promising neuroprotective and anti-inflammatory properties. Among the 14 synthesized compounds, ZA3-ZA5, ZB2-ZB6, and intermediate S5 showed significant anti-neuroinflammatory properties through inhibition of nitric oxide (NO) and tumor necrosis factor-α (TNF-α) production in LPS-stimulated human microglia cells. From 14 compounds, six (ZA2 to ZA6 and intermediate S5) exhibited promising neuroprotective activity by reduced expression of the endoplasmic reticulum (ER) chaperone, BIP, and apoptosis marker cleaved caspase-3 in human neuronal cells. Also, a molecular docking study showed that lead compounds have favorable interaction with active residues of ATF4 and NF-kB proteins. CONCLUSION: The possible mechanism of action was observed through the inhibition of ER stress, apoptosis, and the NF-kB inflammatory pathway. Thus, our study strongly indicates that the novel scaffolds of triazole-pyrimidine-based compounds can potentially be developed as neuroprotective and anti-neuroinflammatory agents.

Crosstalk of oxidative stress, inflammation, apoptosis, and autophagy under reactive oxygen stress involved in difenoconazole-induced kidney damage in carp.

Difenoconazole is a commonly used triazole fungicide in agricultural production. Because of its slow degradation and easy accumulation in the environment, it seriously endangers both animal health and the ecological environment. Therefore, it is hoped that the effects on carp kidneys can be studied by simulating difenoconazole residues in the environment. The experiment was designed with two doses (0.488 mg/L, 1.953 mg/L) as exposure concentrations of difenoconazole for 4 d. Histopathological results showed that difenoconazole could cause severe damage to the kidney structure and extensive inflammatory cell infiltration in carp. Elevated levels of Creatinine, and BUN suggested the development of kidney damage. The DHE fluorescence probe's result suggested that difenoconazole might cause reactive oxygen species (ROS) to accumulate in the kidney of carp. Difenoconazole was found to increase MDA levels while decreasing the activities of CAT, SOD, and GSH-PX, according to biochemical indicators. In addition, difenoconazole could up-regulate the transcription levels of inflammatory factors tnf-α, il-6, il-1ß, and inos. At the same time, it inhibited the transcription level of il-10 and tgf-ß1. The TUNEL test clearly showed that difenoconazole induced apoptosis in the kidney and vastly raised the transcript levels of apoptosis-related genes p53, caspase9, caspase3, and bax while inhibiting the expression of Bcl-2, fas, capsase8. Additionally, TEM imaging showed that clearly autophagic lysosomes and autophagosomes were formed. Elevated levels of LC3II protein expression, increased transcript levels of the autophagy-related gene atg5 as well as decreased transcript levels of p62 represented the generation of autophagy. In conclusion, the study illustrated that oxidative stress, inflammation, apoptosis, and autophagy all played roles in difenoconazole-induced kidney injury in carp, which was closely linked to ROS production. This work provides a valuable reference for studying the toxicity of difenoconazole to aquatic organisms.

Maternal exposure to chiral triazole fungicide tebuconazole induces enantioselective thyroid disruption in zebrafish offspring.

Pesticides could induce long-term impacts on aquatic ecosystem via transgenerational toxicity. However, for many chiral pesticides, the potential enantioselectivity of transgenerational toxicity has yet to be fully understood. In this study, we used zebrafish as models to evaluate the maternal transfer risk of tebuconazole (TEB), which is a chiral triazole fungicide currently used worldwide and has been frequently detected in surface waters. After 28-day food exposure (20 and 400 ng/g) to the two enantiomers of TEB (S- and R-TEB) in adult female zebrafish (F0), increased malformation rate and decreased swimming speed were found in F1 larvae, with R-TEB showing higher impacts than S-enantiomer. Additionally, enantioselective effects on the secretion of thyroid hormones (THs) and expression of TH-related key genes along the hypothalamic-pituitary-thyroid (HPT) axis were found in both F0 and F1 after maternal exposure. Both the two enantiomers significantly disrupted the triiodothyronine (T3) and thyroxine (T4) contents in F0 with different degrees, whereas in F1, significant effects were only found in R-TEB groups with decreasing of both T3 and T4 contents. Most of the HPT axis related genes in F0 were upregulated by TEB and more sensitive to R-TEB than to S-TEB. In contrast, most of the genes in F1 were downregulated by both R- and S-TEB, especially the genes that are primarily responsible for thyroid development and growth (Nkx2-1), TH synthesis (NIS and TSHꞵ) and metabolism (Deio1). Findings from this study highlight the key role of enantioselectivity in the ecological risk assessment of chiral pesticides through maternal transfer.

Sex-specific metabolic dysregulation in digestive glands of green mussels following exposure to triazophos.

As a ubiquitous environmental pollutant in China, triazophos (TP) is known to have neurotoxicity, oxidative stress, and reproductive toxicity to mussels. To investigate the molecular mechanisms of TP toxicity, metabolic changes in the digestive glands of Perna viridis in different sexes were examined after treated with 35 µg/L TP. Notably, 158 significant different metabolites (SDMs) were detected in TP-treated mussels and more than half of the SDMs were lipids and lipid-like molecules, which suggested that TP disturbed the lipid metabolism of P. viridis. In addition, metabolites associated with neurotoxicity and reproductive disturbance were also detected in female and male mussels. Moreover, a larger number of SDMs were found in male mussels (120 SDMs) than females (99 SDMs), and 60 common metabolites exhibited consistent variation tendency and similar magnitude in both sexes. The metabolic alternations in female and male mussels displayed similar protective mechanisms and also sex-specific responses, male mussels were more sensitive to TP exposure. This research provided new data about the molecular mechanisms of TP toxicity and the gender specific changes in mussels after treated by chemicals.

Enantioselective Toxic Effects of Prothioconazole toward Scenedesmus obliquus.

Prothioconazole (PTC) is a broad-spectrum triazole fungicide with one asymmetric center and consists of two enantiomers, R-(-)-PTC and S-(+)-PTC. To address the concern of its environmental safety, the enantioselective toxic effects of PTC on Scendesmus obliquus (S. obliquus) were investigated. PTC racemates (Rac-PTC) and enantiomers exhibited dose-dependent acute toxicity effects against S. obliquus at a concentration from 1 to 10 mg·L-1. The 72 h-EC50 value of Rac-, R-(-)-, and S-(+)-PTC is 8.15, 16.53, and 7.85 mg·L-1, respectively. The growth ratios and photosynthetic pigment contents of the R-(-)-PTC treatment groups were higher than the Rac- and S-(+)-PTC treatment groups. Both catalase (CAT) activities and esterase activities were inhibited in the Rac- and S-(+)-PTC treatment groups at high concentrations of 5 and 10 mg·L-1, and the levels of malondialdehyde (MDA) were elevated, which exceeded the levels in algal cells for the R-(-)-PTC treatment groups. PTC could disrupt the cell morphology of S. obliquus and induce cell membrane damage, following the order of S-(+)-PTC ≈ Rac-PTC > R-(-)-PTC. The enantioselective toxic effects of PTC on S. obliquus provide essential information for its ecological risk assessment.

Characterization of a triazole scaffold compound as an inhibitor of Mycobacterium tuberculosis imidazoleglycerol-phosphate dehydratase.

Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis (TB), employs ten enzymes including imidazoleglycerol-phosphate dehydratase (IGPD) for de novo biosynthesis of histidine. The absence of histidine-biosynthesis in humans combined with its essentiality for Mtb makes the enzymes of this pathway major anti-TB drug targets. We explored the inhibitory potential of a small molecule ß-(1,2,4-Triazole-3-yl)-DL-alanine (DLA) against Mtb IGPD. DLA exhibits an in vitro inhibitory efficacy in the lower micromolar range. Higher-resolution crystal structures of native and substrate-bound Mtb IGPD provided additional structural features of this important drug target. Crystal structure of IGPD-DLA complex at a resolution of 1.75 Å, confirmed that DLA locks down the function of the enzyme by binding in the active site pocket of the IGPD mimicking the substrate-binding mode to a high degree. In our biochemical study, DLA showed an efficient inhibition of Mtb IGPD. Furthermore, DLA also showed bactericidal activity against Mtb and Mycobacterium smegmatis and inhibited their growth in respective culture medium. Importantly, owing to the favorable ADME and physicochemical properties, it serves as an important lead molecule for further derivatizations.

Transcriptomic analysis of Vigna radiata in response to chilling stress and uniconazole application.

BACKGROUND: Chilling injury of mung bean (Vigna radiata (L.)) during the blooming and podding stages is a major agricultural threat in Northeast China. Uniconazole (UNZ) can alleviate water deficit stress in soybean and waterlogging stress in mung bean. However, there has been no report on the effect of UNZ application on the growth and transcriptomic profile of mung bean under chilling stress. RESULTS: UNZ application before chilling stress at the R1 stage alleviated the decline in mung bean yield. UNZ delayed the decrease in leaf chlorophyll content under chilling stress at the R1 stage and accelerated the increase in leaf chlorophyll content during the recovery period. Eighteen separate RNA-Seq libraries were generated from RNA samples collected from leaves exposed to six different treatment schemes. The numbers of DEGs specific for UNZ treatment between D1 + S vs. D1 and D4 + S vs. D4 were 708 and 810, respectively. GO annotations showed that photosynthesis genes were obviously enriched among the genes affected by chilling stress and UNZ application. KEGG pathway enrichment analysis indicated that 4 pathways (cutin, suberin and wax biosynthesis; photosynthesis; porphyrin and chlorophyll metabolism; and ribosome) were downregulated, while plant-pathogen interaction was upregulated, by chilling stress. UNZ application effectively prevented the further downregulation of the gene expression of members of these 4 KEGG pathways under chilling stress. CONCLUSIONS: UNZ application effectively delayed the decrease in photosynthetic pigment content under chilling stress and accelerated the increase in photosynthetic pigment content during the recovery period, thus effectively limiting the decline in mung bean yield. UNZ application effectively prevented the further downregulation of the gene expression of members of 4 KEGG pathways under chilling stress and increased mung bean tolerance to chilling stress.

Designing bacterial consortia for the complete biodegradation of insensitive munitions compounds in waste streams.

Insensitive munitions compounds (IMCs), such as 2,4-dinitroanisole (DNAN) and 3-nitro-1,2,4-triazol-5-one (NTO), are replacing conventional explosives in munitions formulations. Manufacture and use of IMCs generate waste streams in manufacturing plants and load/assemble/pack facilities. There is a lack of practical experience in executing biodegradation strategies to treat IMCs waste streams. This study establishes a proof-of-concept that bacterial consortia can be designed to mineralize IMCs and co-occurring nitroaromatics in waste streams. First, DNAN, 4-nitroanisole (4-NA), and 4-chloronitrobenzene (4-CNB) in a synthetic DNAN-manufacturing waste stream were biodegraded using an aerobic fluidized-bed reactor (FBR) inoculated with Nocardioides sp. JS 1661 (DNAN degrader), Rhodococcus sp. JS 3073 (4-NA degrader), and Comamonadaceae sp. LW1 (4-CNB degrader). No biodegradation was detected when the FBR was operated under anoxic conditions. Second, DNAN and NTO were biodegraded in a synthetic load/assemble/pack waste stream during a sequential treatment comprising: (i) aerobic DNAN biodegradation in the FBR; (ii) anaerobic NTO biotransformation to 3-amino-1,2,4-triazol-5-one (ATO) by an NTO-respiring enrichment; and (iii) aerobic ATO mineralization by an ATO-oxidizing enrichment. Complete biodegradation relied on switching redox conditions. The results provide the basis for designing consortia to treat mixtures of IMCs and related waste products by incorporating microbes with the required catabolic capabilities.

Widening the Lens on Prothioconazole and Its Metabolite Prothioconazole-Desthio: Aryl Hydrocarbon Receptor-Mediated Reproductive Disorders through in Vivo, in Vitro, and in Silico Studies.

Reproductive disorders are a serious public health problem worldwide. Epidemiological data suggest that exposure to environmental pollutants is associated with the onset of reproductive disorders. However, the effects in reproductive health and exact mechanism of action of representative agricultural compounds prothioconazole (PTC) and its metabolite prothioconazole-desthio (dPTC) on mammals remain unclear. Here, we studied the physiological effects of the exposure to environmentally relevant doses of PTC and dPTC in mice reproductive systems. Combining in vivo, in vitro, and in silico studies, we observed that PTC and dPTC disrupt reproductive health by inducing metabolic perturbation, induction of apoptosis, and inflammation in gonadal tissue, which are achieved via activation of the aryl hydrocarbon receptor (AhR). Convincingly, the addition of alternate-day injections of CH223191 (an AhR inhibitor) to the 30-day exposure regimen ameliorated ovarian tissue damage, as evidenced by decreased TUNEL-positive cells and partially restored the inflammation and apoptotic factor levels. This study comprehensively reports the toxic effects of low-dose PTC and dPTC in the reproductive system in vivo and identifies AhR as a potential therapeutic target for the amelioration of reproductive disorders caused by similar endocrine-disrupting chemicals.

Paclobutrazol Ameliorates Low-Light-Induced Damage by Improving Photosynthesis, Antioxidant Defense System, and Regulating Hormone Levels in Tall Fescue.

Paclobutrazol (PBZ) is a plant-growth regulator (PGR) in the triazole family that enhances plant tolerance to environmental stresses. Low-light (LL) intensity is a critical factor adversely affecting the growth of tall fescue (Festuca arundinacea Schreb.). Therefore, in this study, tall fescue seedlings were treated with PBZ under control and LL conditions to investigate the effects of PBZ on enhancing LL stress resistance by regulating the growth, photosynthesis, oxidative defense, and hormone levels. Our results reveal that LL stress reduced the total biomass, chlorophyll (Chl) content, photosynthetic capacity, and photochemical efficiency of photosystem II (PSII) but increased the membrane lipid peroxidation level and reactive oxygen species (ROS) generation. However, the application of PBZ increased the photosynthetic pigment contents, net photosynthetic rate (Pn), maximum quantum yield of PSII photochemistry (Fv/Fm), ribulose-1,5-bisphosphate carboxylase (RuBisCO) activity, and starch content. In addition, PBZ treatment activated the antioxidant enzyme activities, antioxidants contents, ascorbate acid-glutathione (AsA-GSH) cycle, and related gene expression, lessening the ROS burst (H2O2 and O2∙-). However, the gibberellic acid (GA) anabolism was remarkably decreased by PBZ treatment under LL stress, downregulating the transcript levels of kaurene oxidase (KO), kaurenoic acid oxidase (KAO), and GA 20-oxidases (GA20ox). At the same time, PBZ treatment up-regulated 9-cis-epoxycarotenoid dioxygenase (NCED) gene expression, significantly increasing the endogenous abscisic acid (ABA) concentration under LL stress. Thus, our study revealed that PBZ improves the antioxidation and photosynthetic capacity, meanwhile increasing the ABA concentration and decreasing GA concentration, which ultimately enhances the LL stress tolerance in tall fescue.

Novel Hybrid 1,2,4- and 1,2,3-Triazoles Targeting Mycobacterium Tuberculosis Enoyl Acyl Carrier Protein Reductase (InhA): Design, Synthesis, and Molecular Docking.

Tuberculosis (TB) caused by Mycobacterium tuberculosis is still a serious public health concern around the world. More treatment strategies or more specific molecular targets have been sought by researchers. One of the most important targets is M. tuberculosis' enoyl-acyl carrier protein reductase InhA which is considered a promising, well-studied target for anti-tuberculosis medication development. Our team has made it a goal to find new lead structures that could be useful in the creation of new antitubercular drugs. In this study, a new class of 1,2,3- and 1,2,4-triazole hybrid compounds was prepared. Click synthesis was used to afford 1,2,3-triazoles scaffold linked to 1,2,4-triazole by fixable mercaptomethylene linker. The new prepared compounds have been characterized by different spectroscopic tools. The designed compounds were tested in vitro against the InhA enzyme. At 10 nM, the inhibitors 5b, 5c, 7c, 7d, 7e, and 7f successfully and totally (100%) inhibited the InhA enzyme. The IC50 values were calculated using different concentrations. With IC50 values of 0.074 and 0.13 nM, 7c and 7e were the most promising InhA inhibitors. Furthermore, a molecular docking investigation was carried out to support antitubercular activity as well as to analyze the binding manner of the screened compounds with the target InhA enzyme's binding site.

Repurposing of FDA-approved drugs against active site and potential allosteric drug-binding sites of COVID-19 main protease.

The novel coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still has serious negative effects on health, social life, and economics. Recently, vaccines from various companies have been urgently approved to control SARS-CoV-2 infections. However, any specific antiviral drug has not been confirmed so far for regular treatment. An important target is the main protease (Mpro ), which plays a major role in replication of the virus. In this study, Gaussian and residue network models are employed to reveal two distinct potential allosteric sites on Mpro that can be evaluated as drug targets besides the active site. Then, Food and Drug Administration (FDA)-approved drugs are docked to three distinct sites with flexible docking using AutoDock Vina to identify potential drug candidates. Fourteen best molecule hits for the active site of Mpro are determined. Six of these also exhibit high docking scores for the potential allosteric regions. Full-atom molecular dynamics simulations with MM-GBSA method indicate that compounds docked to active and potential allosteric sites form stable interactions with high binding free energy (∆Gbind ) values. ∆Gbind values reach -52.06 kcal/mol for the active site, -51.08 kcal/mol for the potential allosteric site 1, and - 42.93 kcal/mol for the potential allosteric site 2. Energy decomposition calculations per residue elucidate key binding residues stabilizing the ligands that can further serve to design pharmacophores. This systematic and efficient computational analysis successfully determines ivermectine, diosmin, and selinexor currently subjected to clinical trials, and further proposes bromocriptine, elbasvir as Mpro inhibitor candidates to be evaluated against SARS-CoV-2 infections.